Glycosome turnover inLeishmania majoris mediated by autophagy

Abstract

Abbreviations: ATG, autophagy-related; GFP, green fluorescent protein; mC, mCherry fluorescent protein; MVT, multivesicular tubule; RFP, red fluorescent protein; TEM, transmission electron microscopy.Autophagy is a central process behind the cellular remodeling that occurs during differentiation of Leishmania, yet the cargo of the protozoan parasite's autophagosome is unknown. We have identified glycosomes, peroxisome-like organelles that uniquely compartmentalize glycolytic and other metabolic enzymes in Leishmania and other kinetoplastid parasitic protozoa, as autophagosome cargo. It has been proposed that the number of glycosomes and their content change during the Leishmania life cycle as a key adaptation to the different environments encountered. Quantification of RFP-SQL-labeled glycosomes showed that promastigotes of L. major possess »20 glycosomes per cell, whereas amastigotes contain »10. Glycosome numbers were significantly greater in promastigotes and amastigotes of autophagy-defective L. major Datg5 mutants, implicating autophagy in glycosome homeostasis and providing a partial explanation for the previously observed growth and virulence defects of these mutants. Use of GFP-ATG8 to label autophagosomes showed glycosomes to be cargo in »15% of them; glycosome-containing autophagosomes were trafficked to the lysosome for degradation. The number of autophagosomes increased 10-fold during differentiation, yet the percentage of glycosome-containing autophagosomes remained constant. This indicates that increased turnover of glycosomes was due to an overall increase in autophagy, rather than an upregulation of autophagosomes containing this cargo. Mitophagy of the single mitochondrion was not observed in L. major during normal growth or differentiation; however, mitochondrial remnants resulting from stress-induced fragmentation colocalized with autophagosomes and lysosomes, indicating that autophagy is used to recycle these damaged organelles. These data show that autophagy in Leishmania has a central role not only in maintaining cellular homeostasis and recycling damaged organelles but crucially in the adaptation to environmental change through the turnover of glycosomes.

The Transcriptome ofLeishmania majorDevelopmental Stages in Their Natural Sand Fly Vector

“…In L. major , autophagy was shown to be essential for metacyclogenesis and was significantly upregulated in culture metacyclics (55, 56). It was shown in a later study that recycling of the glycosome compartment via autophagy is an important aspect of this response (57). ATG8 is involved in completion and expansion of the autophagosome vesicles and was previously used as a marker for autophagy in Leishmania (56–58).…”

“…It was shown in a later study that recycling of the glycosome compartment via autophagy is an important aspect of this response (57). ATG8 is involved in completion and expansion of the autophagosome vesicles and was previously used as a marker for autophagy in Leishmania (56–58). In our study, putative ATG8 mRNAs on chromosomes 9 and 19 were among the most strongly upregulated in the NP stage (Fig.…”

ATP-driven and AMPK-independent autophagy in an early branching eukaryotic parasite

“…21 Additionally, taking advantage of the molecular genetic accessibility of T. brucei, a role of autophagy in cell death and glycosome-turnover and differentiation has been proposed. 20,[22][23][24] The prodeath role of autophagy is also reported in Toxoplasma gondii and Plasmodium berghei. 25,26 Trypanosoma brucei transmits between mammal hosts via tsetse flies.…”

Structure, Properties, and Function of Glycosomes in Trypanosoma cruzi

“…The mechanism by which the life-cycle stage dependent glycosome reprogramming occurs has not yet been studied in T. cruzi, but has been addressed in T. brucei and Leishmania spp. (Hart et al, 1984;Mottram and Coombs, 1985;Colasante et al, 2006;Herman et al, 2008;Vertommen et al, 2008;Brennand et al, 2011;Cull et al, 2014). Data indicate that this reprogramming involves mainly degradation of organelles containing the redundant enzyme repertoire by pexophagy-i.e., the autophagy process characteristic for peroxisome degradation-while new organelles are formed with an enzyme content that is appropriate for the conditions to be encountered by the new developmental form.…”